Human infection by viruses is one of the major health problems facing the mankind. For example, about 180 million people worldwide are infected with hepatitis C virus (HCV). Although an array of pharmaceutical drugs are available now, the use of these drugs requires the monitoring of the efficacy of the drugs by measuring the virus concentration in blood samples, commonly known as the viral load. The viral load is normally determined by measuring the viral nucleic acid concentration in the blood samples (serum or plasma). A number of methods have been developed for measuring viral nucleic acid concentration with the most commonly used method being amplification methods, such as the polymerase chain reaction (PCR).
The most cumbersome step to carry out a PCR is the isolation of viral nucleic acid from the sample, particularly for those samples with very low viral concentration. For the detection of low viral load, the samples are often centrifuged to enrich the viruses using an ultracentrifugation process, which requires an expensive ultracentrifuge and a long centrifugation time. An alternative method for enriching viral particles is binding the virus particles to a solid phase. Thereby, the virus particles are enriched from the original sample.
The viral nucleic acids can then be isolated from the virus particle enriched fraction. Thereby, advantageously, a concentration of the viral particles from large sample volumes is possible.
However, the amount and recoverability of available virus particles and the corresponding viral nucleic acids can decrease substantially over a period of time due to degradation. Thus, the virus load decreases over time, thereby potentially falsifying the results. However, a successful medical treatment of infectious diseases like HCV depends largely on the reliable and accurate quantification of the viral load and hence the viral nucleic acids in patient samples. But clinics where blood collection takes place and laboratories performing viral level determination are often physically separated. Delivering samples from clinics to laboratories often takes place under unfavourable conditions like temperature variations, mechanical shocks and delay resulting in a decrease of viral level loads. In order to stabilize the viral nucleic acid levels in whole blood samples through shipment a blood collection tube containing a stabilizing reagent is required.
Blood samples containing virus particles are usually collected in blood collection tubes containing spray-dried or liquid EDTA as anticoagulant (e.g. BD Vacutainer K2EDTA). EDTA chelates magnesium, calcium and other bivalent metal ions, thereby inhibiting enzymatic reactions, such as e.g. blood clotting or DNA degradation due to DNases. However, EDTA does not stabilize virus particles which usually consist of two or three parts: the genetic material made from either DNA or RNA, long molecules that carry genetic information; a protein coat that protects these genes; and in some embodiments an envelope of lipids that surrounds the protein coat when they are outside a cell. Thus, viruses as well as viral nucleic acids are susceptible to degradation.
Thus, it is inter alia an object of the present invention to provide a method for stabilizing virus particles in a biological sample, in particular a sample obtained from a human body, and to provide a suitable method for isolating virus particles and/or viral nucleic acids from a stabilized sample.